ABSTRACT The majority of breast cancer cases are estrogen receptor (ER?)-positive. While hormonal therapy improves clinical outcomes for about half of patients with ER?-positive breast cancer, de novo or acquired resistance represents a significant clinical challenge. Among several underlying mechanisms, hot-spot point mutants of ER? are known to confer therapeutic resistance due to their estrogen-independent transcriptional activity. Thus, mitigating aberrant transcription activity of these ER? mutants holds promise for overcoming therapeutic resistance in treatment of ER?-positive breast cancer. As the second member of the ER family, ER? is capable of interfering with ER? activity through heterodimerization and/or competing for common chromatin binding sites. This ER?-interfering function of ER? could be utilized to overcome the activity of therapeutically resistant ER? mutants. However, clinical feasibility of this approach is vastly under-explored, as little is known about how ER?-interfering activity of ER? is mobilized. Our preliminary work discovered a functionally important phosphotyrosine switch in ER?. Specifically, we found that unphosphorylated ER? is particularly potent in heterodimerization and functional interference with ER?. We therefore hypothesize that unphosphorylated ER? in ER?-positive breast cancer can help overcome ER? mutant-mediated therapeutic resistance. We further envision that pharmacological agents that fine-tune the phosphotyrosine status of ER? could be clinically useful in stimulating its ER?-interfering activity. We will test this novel hypothesis through two Specific Aims. First, we will use in vitro and patient- derived xenograft models to determine the impact of ER? phosphorylation status on ER?-mediated therapeutic resistance. Second, we will use molecular and pharmacological tools to elucidate the mechanism by which ER? phosphorylation status regulates the ER?/? crosstalk. The concept of overcoming therapeutic resistance by rallying a particular form of ER? represents a novel concept. Furthermore, because the inhibitor of the upstream kinase for the phosphotyrosine switch is clinically available and ER?-specific agonists are well tolerated in humans, our work provides multiple druggable targets for fine-tuning ER? activities and imminent translatability for treating ER?-positive breast cancer. Our proposed study promises both conceptual and translational advances in understanding of how to overcome therapeutic resistance to hormonal therapy, a pressing clinical challenge in breast cancer treatment.